Amazing stuff!
"The virus-replicating machinery of the SARS-Cov-2 virus and the ribonucleic acid (RNA) molecules it produces as it replicates are located in physically distinct areas of an infected cell. This observation, made by researchers at Stanford University in the US using a new multicolour super-resolution microscopy imaging technique, provides fresh insights into the life cycle of coronaviruses and could aid the development of new therapeutics to fight them. ...
studied coronavirus infection at the nanoscale using high-throughput confocal and super-resolution microscopy. The researchers used a less harmful human coronavirus, HCoV-229E, as their model, because it is similar to SARS-Cov-2 in that it contains an envelope studded with protein spikes surrounding a strand of gRNA. These proteins include those that make gRNA copies and those that assemble into the “packaging” that wraps around the gRNA to make new viruses.
Differently coloured fluorescent tags
In their study ... focused on gRNA and another type of RNA, double-stranded RNA (dsRNA), which makes new copies of the virus. They began by labelling the gRNA and dsRNA with differently coloured fluorescent tags – magenta for the gRNA and green for the dsRNA – so that they could study where the two types of RNA migrated in a cell. The technique they used to do this, confocal fluorescence microscopy, is good at recording light emitted from the fluorescent labels. However, its spatial resolution is limited to imaging structures 250 nm in size or bigger. This is a problem because coronaviruses are half this size, and the proteins and RNA they contain are smaller still, at 50 nm and 10 nm or less. ... To overcome this problem, the researchers turned to super-resolution fluorescence microscopy, which brings these structures into sharper focus, allowing objects as small as 10 nm across to be imaged. This is well below the optical diffraction limit. ..."
From the abstract:
"The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the third human coronavirus within 20 years that gave rise to a life-threatening disease and the first to reach pandemic spread. ... Here, we present a robust and generalizable framework combining high-throughput confocal and super-resolution microscopy imaging to study coronavirus infection at the nanoscale. Using the model human coronavirus HCoV-229E, we specifically labeled coronavirus genomic RNA (gRNA) and double-stranded RNA (dsRNA) via multi-color RNA immunoFISH and visualized their localization patterns within the cell. The 10-nm resolution achieved by our approach uncovers a striking spatial organization of gRNA and dsRNA into three distinct structures and enables quantitative characterization of the status of the infection after antiviral drug treatment. ..."
Multi-color super-resolution imaging to study human coronavirus RNA during cellular infection (open access)
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